Absence of a dnaspecific signal was observed for the majority of. Xav939, a tankyrase 1 inhibitior, promotes cell apoptosis in. Once the culture is established, freeze back a portion of the material, and. Retinoic acid reduces human neuroblastoma cell migration. The karyotype is hyperdiploid female with a modal chromosome number of 47. Identification and classification of genes regulated by. Just follow the protocol for the size of your plates, but before transfecting split the shsy5y into plates coated with fibronectin at 1. Fectopro transfection reagent for bioproduction polyplus. From cell lines to pluripotent stem cells for modelling. This information is derived based on pharmacological data in reference to a doseresponse curve. Maintaining healthy cells is the key to experimental success and reproducible research results. The parental cell line was generated in 1970 from a bone marrow biopsy that contains both neuroblastlike and epitheliallike cells 15. A protocol including 15% fbs in the culture condition has been previously described as a method for sequential treatment of shsy5y cells with ra and bdnf, although the present study used a dmemf12 1. Complete growth medium supplemented with 5% vv dmso.
Xav939, a tankyrase 1 inhibitior, promotes cell apoptosis. These are the protocols that were used to produce the cells that we have sent to you. In vitro and in vivo efficacy of nonpsychoactive cannabidiol. Sh sy5y cells have a reported saturation density greater than 1 x 10 6 cellscm 2. The establishment of a reliable cytotoxic system with sknsh. Karyotype the cell line is hyperdiploid human female xx, with the modal chromosome number of 47. Mar 16, 2016 the human nbl cell lines sknsh 25 and imr32 26 were purchased from atcc manassas, va, u. Briefly, random cultured cells from two clone subtypes of shsy5y and sknsh were seeded. Of note, sknbe2c is a subclone derived from the parental sknbe2 cell line 12 and shsy5y was derived from the shsy subclone of the parental sknsh cell line.
Mouse neuronal stem cells nscs from ventral mesencephalon were cultured and differentiated following the manufacturers protocol stemcell technologies. Human brain neuroepithelioma cells sknmc can be ordered from atcc as a frozen ampoule with 2 x 106 per 1ml volume. I am currently trying to grow sknsh cells and they are growing extremely slowly, much slower than any other cell line i have ever used. Volumes used in this protocol are for 75 cm 2 flask. Sknas are distributed for research purposes only by the memorial sloankettering cancer center. A subline of the neuroblastoma sk n sh line derived from a metastatic bone tumor. Cell culture is the process by which cells are grown under controlled conditions, generally outside their natural environment.
Furthermore, it was noted that earlier passages and some sublines of skn. It has also been used to study the potential mechanisms and domain requirements of the hhsj1a dnaj heat shock protein family member b2 mediated reduction in superoxide dismutase 1 sod1 aggregation. It is now widely regarded as having originated from an askins tumor ewing family of tumors. The original cell line sk n sh presents two phenotypes characterized by different morphological and biochemical features. Neuroblastoma cell lines center for cancer research.
Amounts of dna and volumes of reagent fectopro and competitor for transfection according to manufacturers recommendations. Receipt of frozen cells and starting cell cultures. Morphologic conversion of a neuroblastomaderived cell. Every step of the way, a wide range of cell health products maintaining healthy cells is the key to experimental success and reproducible research results.
Morroniside protects sknsh human neuroblastoma cells against. Biedler and is different from sknmc as it displays higher levels of dopamine betahydroxylase as well as a longer doubling time. Accurate rtqpcr gene expression analysis on cell culture lysates. Sh sy5y neuroblastlike cells are a subclone of the parental neuroblastoma cell line sk n sh. Sk n sh reverse transfection protocol 24well plate. These cells harbor the oncogenic ewsfli1 chromosomal rearrangement. These cells are typically locked in an early neuronal differentiation stage, characterized biochemically by the low presence of neuronal markers biedler et al. Htb11 is a neuroblastoma cell line derived from human bone marrow. Sk n sh standard transfection protocol 24well plate. Differentiation of the shsy5y human neuroblastoma cell line. The 2cq method was used for the relative quantification of the gene.
These vessels are seeded with cells, incubated to ensure cell growth and then filled completely with medium for shipping. Sk n mc cells exhibit an epithelial cell type and are neuroblastoma cells, which makes them useful for research related to brain cancers. Cell culture protocol for shsy5y neuroblastoma cells. Shifra ash, schneider childrens medical center of israel 28. Recommendations are based on inhouse testing by the transfection experts. Sh cell line has been used in dark toxicity testing of a chloroindiumphthalocyan ine photosensitizer inpc. Shsy5y cell transfection cell biology protocol online. Recommended transfection protocols for 24well plate. Cell culture protocol for shsy5y neuroblastoma cells human. Epigenomic profiling of neuroblastoma cell lines scientific.
Examine the cell cultures after 24 hours and subculture as needed. This growth formation can be referred to as the overeasy formation sh sy5y is a human derived cell line used in scientific research. Shsy5y cells have a stable karyotype consisting of 47 chromosomes, and can be differentiated from a neuroblastlike. Shsy5y atcc crl2266 homo sapiens bone marrow neuroblast. To give you confidence in the health of your cells every step of the way, weve highlighted the technologies and products within cell biology that are critical to maintaining optimal cell. Tnks1 in shsy5y and sknsh cells was inhibited by xav939 of 1 and 0. Tested on various primary cells and cell lines, jetoptimus proved its superiority by reaching higher transfection efficiencies and gene expression than main.
Shsy5y cells are often used as in vitro models of neuronal function and differentiation. Comparison of neurons derived from mouse p19, rat pc12. Shsy5y are a subclone of the sknsh neuroblastoma cells, obtained in 1978 starting from a 4yearold girl bone marrow biopsy biedler et al. Shsy5y cells were derived through three successive subclones of the parental sknsh cell line1. We isolated two clones 1 and 2 and named this cell line as sknsh. Jan, 2012 to evaluate the performance of dnase treatment on crude cell lysates, single replicate cellstoct lysates were prepared both in the presence and the absence of dnase i in the lysis buffer for four neuroblastoma cell lines sh ep, sk n as, ngp, imr32. Sh sy5y cells have a stable karyotype consisting of 47 chromosomes, and can be differentiated from a neuroblastlike. Temozolomide resistance in glioblastoma cell lines. Sknmc cells exhibit an epithelial cell type and are neuroblastoma cells, which makes them useful for research related to brain cancers. Nsh neuroblastoma cell line, mutated nras genes are not commonly detected in human neuroblastoma tumors 11533 and mutations in nras, kiras and hras were not detected in another series 024. Cell culture neuroblastoma cell lines shep, sknas, ngp, imr32 were cultured in 96well culture plates at a density of 10,000 cellswell. The sknmc neuroblastoma cell line has a higher cholinergic activity relative to other nb cell lines, and this coupled with its tumorigenicity make it a powerful research tool. Transitx2tm dynamic delivery system mirus bio transit.
Dorschner ordering information sk n sh can be ordered from atcc as a frozen ampule. We recommend that you read through these protocols prior to thawing your cells, and follow them until you have established your own cell bank of frozen vials. Sh sy5y cells can form mounds of undifferentiated cells, which then spread differentiated cells into the surrounding area. The parental cell line was generated in 1970 from a bone marrow biopsy that contains both neuroblast. Description sk n sh is a neuroblastoma cell line that displays epithelial morphology and grows in adherent culture. Half maximal inhibitory concentration, or ic 50, is a measurement representing the halfway point in which a compound of interest produces complete inhibition of a biological or biochemical function. In this context, the popular shsy5y neuroblastoma cell line and its use in in vitro systems is described. When stimulated with retinoic acid ra, shsy5y differentiates to dopaminergic neuronlike cells 9. To perform the retinoic acid resistance screen in sknsh cells, vbimsd1, sd2 and sd3. Also, sknsh is used in cellmediated cytotoxicity assays as a target cell.
In this normalization method, values are transformed to values. The respective cell typespecific optimized protocol or recommendation is available under instructions or in our knowledge database. Treatment with alltransretinoic acid causes these cells to differentiate and adopt a neuronal phenotype, characterized by extensive neurite outgrowth. Reduction of mir21 induces sknsh cell apoptosis and inhibits. Sknas transfection kit neuroblastoma cells altogen. Furthermore, it was noted that earlier passages and some sublines of sk n sh didnt have a mutated n ras. Cell culture growth conditions,protein expression,stable cell transfection,transient transfection. To investigate this mechanism, we compared the gene expression profiles in sk n sh cells and two subtypes of sh sy5y cells sh sy5ya and sh sy5ye, each of which show a. Feb 17, 2016 sh sy5y neuroblastlike cells are a subclone of the parental neuroblastoma cell line sk n sh. To give you confidence in the health of your cells every step of the way, weve highlighted the technologies and products within cell biology that are critical to maintaining optimal cell health. Pdf differentiation of the shsy5y human neuroblastoma cell. A subline of the neuroblastoma sknsh line derived from a metastatic bone tumor. Transcriptomic profiling of 39 commonlyused neuroblastoma. The sk n mc neuroblastoma cell line has a higher cholinergic activity relative to other nb cell lines, and this coupled with its tumorigenicity make it a powerful research tool.
This makes them particularly useful for delineating signaling pathways involved in neuronal differentiation. Request pdf the establishment of a reliable cytotoxic system with sknsh neuroblastoma cell culture a reliable in vitro cytotoxic system is essential in. This cell line is a human catecholaminergic neuroblastoma derived from sknsh, which resembles immature sympathetic neuroblasts in culture biedler et al. Jan 24, 2017 an in vitro model widely used in pd research is the neuroblastoma sh sy5y cell line. In the sknsh cell line, tmz treatment caused cells to appear in the g2m phase 21. Protocol here, which will probably recommend a product, seeing as it is a sigma protocol.
Multiple neurotransmitter synthesis by human neuroblastoma cell lines and clones. Cell line nucleofector tm kit v lonza bioscience lonza. These conditions vary for each cell type, but generally consist of a suitable vessel with a substrate or medium that supplies. They are reported to exhibit moderate levels of dopamine beta hydroxylase activity. The parental cell line was generated in 1970 from a. They were initially found to contain doubleminute chromosomes, which were lost upon prolonged in vitro culture. Cells are differentiated with retinoic acid atra for 48 hours. Accurate rtqpcr gene expression analysis on cell culture.
Comparison between proliferative and neuronlike shsy5y. Description sknsh is a neuroblastoma cell line that displays epithelial morphology and grows in adherent culture. I am currently trying to grow sk n sh cells and they are growing extremely slowly, much slower than any other cell line i have ever used. There are two marker chromosomes and n7 is trisomic.
The human nbl cell lines sknsh 25 and imr32 26 were purchased from atcc manassas, va, u. Shsy5y sknsh skov3 sw480 tc28 a2 tg havsmc t47d thp1 u2 os vero. Transfect at 8090% with fugenehd or at 75% with lipofectamine. P19 cells were isolated in 1981 from a mouse teratocarcinoma 10.
Wicell feeder independent pluripotent stem cell protocols. The human cell line shsy5y is a subclone of sknsh, which was isolated in 1970 from the bone marrow aspiration of a 4yearold girl with metastatic neuroblastoma 8. Dorschner ordering information sknsh can be ordered from atcc as a frozen ampule. The shsy5y cell line in parkinsons disease research. To evaluate the performance of dnase treatment on crude cell lysates, single replicate cellstoct lysates were prepared both in the presence and the absence of dnase i in the lysis buffer for four neuroblastoma cell lines shep, sknas, ngp, imr32.
The cell line is a subclone of the parent cell line sknsh which was originally established from a bone marrow biopsy of a neuroblastoma patient. The remaining cell lines had concordant mycn and myc mrna and protein expression, thus, cogn415, kelly, nb1643, lan5, and ngp were chosen for mycn chipseq while nb69, sknas, and sknsh. Note 2 processing flask cultures some atcc cell, are shipped as growing cultures in culture vessels. They are adrenergic in phenotype but also express dopaminergic markers and, as. Cell growth protocol for htb11 cell line htb11 ucsc genome. The following table lists the optimal transitx2 volume for sirna transfections in the indicated cell types when using 25 nm sirna per well of a 6well plate. Homo sapiens sh sy5y cell line description neuroblastoma clonal subline of the neuroepithelioma cell line sk n sh from the 1970 bone marrow biopsy of a 4yearold girl with metastatic neuroblastoma, trisomy of 1q. Also, fugene hd might not work in serumcontaining medium, i transfect in optimem for 46hr before switching to antibiotic containingserumcontaining medium. Sknmc transfection kit neuroblastoma cells altogen. Sknsh human neuroblastoma cells type culture collection of the. In this optimized protocol the best nucleofection conditions are indicated. The nub6 27 and lan1 cell lines were kindly provided by dr. This line is a subline of the sk n sh cell line, which was established in culture in 1970 from a bone marrow biopsy of a metastatic neuroblastoma of a 4yearold female and has undergone three rounds of clonal selection 6. These cells were also subjected to two tmz cycles in order to observe modulation in the cell cycle distribution.
Sk n sh has been used as a target cell line in cell mediated cytotoxicity assays and is a suitable transfection host. Sh sy5y cells exhibit a neuronal phenotype when treated with alltrans retinoic acid ra, but the molecular mechanism of activation in the signalling pathway mediated by phosphatidylinositol 3kinase pi3k is unclear. Mar 28, 2017 of note, sk n be2c is a subclone derived from the parental sk n be2 cell line 12 and sh sy5y was derived from the sh sy subclone of the parental sk n sh cell line. Shifra ash, schneider childrens medical center of israel 28 sknsh cells were cultured in eagle minimum essential. Description sknmc was originally described as a neuroblastoma cell line. Following inhibition of tnks1 in shsy5y and sknsh cells, protein levels of bcl2 were both reduced p line. Shsy5y neuroblastlike cells are a subclone of the parental neuroblastoma cell line sknsh. Sknsh and pc12 cells were differentiated following atcc protocol.
The cells were allowed to grow for a month until visible colonies were observed, replica plated and genotyped by pcr. The crude cell lysates were directly used as input for qpcr using four dna specific assays. Shsy5y cells have a reported saturation density greater than 1 x 10 6 cellscm 2. The original cell line, called sk n sh, from which it was subcloned was isolated from a. Sknsh transfection kit for neuroblastoma cells, htb11. As you thaw and expand your initial cell vials, we strongly encourage you to establish your own bank of frozen vials as soon as pos sible. Supplementary material and methods cell culture and retroviral. The three human diseases most frequently mentioned in literature for shsy5y were neuroblastoma, alzheimers disease ad, and parkinsons disease pd additional file 1. Appropriately scale the amounts of transitx2 with the surface area of the cell culture vessel please refer to table 2 of the transitx2 protocol. Shsy5y is a human derived cell line used in scientific research. N sh neuroblastoma cell line, mutated n ras genes are not commonly detected in human neuroblastoma tumors 11533 and mutations in n ras, kiras and hras were not detected in another series 024. Originally derived from a metastatic bone tumor biopsy, shsy5y atcc crl2266 cells are a subline of the parental line sknsh atcc htb11. After the cells of interest have been isolated from living tissue, they can subsequently be maintained under carefully controlled conditions. Sknsh, ske6, and skmute6 cells were plated in 60mm 2 tissue culture plates at a cell density of 2 x 10 5 cellsplate.
Systems genomics evaluation of the shsy5y neuroblastoma. Homo sapiens shsy5y cell line description neuroblastoma clonal subline of the neuroepithelioma cell line sknsh from the 1970 bone marrow biopsy of a 4yearold. Furthermore, it was noted that earlier passages and some sublines of sknsh didnt have a mutated nras. Sh sy5y sk n sh sk ov3 sw480 tc28 a2 tg havsmc t47d thp1 u2 os vero.
411 1484 712 1201 574 550 1429 424 450 114 114 1572 1041 621 775 1594 136 1435 1275 1224 1084 215 676 1279 1191 718 1329 1449 291 949 281 1432 853 865 1280 1618 75 1394 952 1363 1203 1136 468 1443 917 1378 1129 755 667 29 1471